This invention relates to the recovery of active forms of proteins which are produced within microbial cultures through recombinant DNA techniques. More specifically, the invention concerns isolation of rennin or a precursor thereof, which is active in milk clotting or proteolysis, from proteins precipitated within bacterial cells as a result of expression of recombinant DNA.
The use of rennin in food processing where clotting of milk is a desirable step has been known for many centuries. Originally, in the process of making cheeses or other milk derived products, strips cut from the stomachs of domesticated young mammals were used as catalysts in the preparations. Later, extracts from, for example, calf stomach mucosa were substituted in cheese making and in preparation of puddings and the like. At the present time, calf stomach remains the major source of these materials, although the processes for extracting it have increased in sophistication.
Recently, it has been possible to prepare several known protein sequences using recombinant DNA techniques by transfecting microbes with gene sequences coding for the desired proteins. Rennin and its precursors are among these. See for example copending application U.S. Pat. Ser. No. 452,227, filed Dec. 22, 1982, now abandoned and British Patent Application GB 2,091,271 A, published 28 Jul. 1982. In the process described in the copending application, the desired rennin protein or its precursor is precipitated within the cell as a "refractile body". The present application describes a process for recovering active rennin activity from the refractile body protein produced in this way.